Vitamin A aerosol-inhalant preparations and method

ABSTRACT

A pharmaceutical preparation consisting of retinoic acid and/or an ester of retinoic acid and/or an ester of retinol as active substances, which are present in the preparation form of aerosol, is advantageously suitable for topical treatment of mucosal diseases in man and animal.

This is a Continuation of application Ser. No. 07/839,547 filed Feb. 19,1992, now abandoned, which was a Division of application Ser. No.07/346,439, filed May 2, 1989, now U.S. Pat. No. 5,112,598.

BACKGROUND OF THE INVENTION

The invention relates to a pharmaceutical preparation.

The term vitamin A stands for a number of chemically similar compoundswith different effect in the human and animal organism. These include asessential groups the alcohol of the vitamin, retinol, the transport formof the vitamin in blood, the aldehyde, the biologically activemetabolite in the visual process, retinal ester, the storage form of thevitamin in the liver and in mucous membranes and reproductive glands,and retinoic acid with various derivatives which play a part inparticular in the differentiation processes of the skin. The vitamin isessential to man, i.e. a vitamin deficiency results if it is not takenin with food. The vitamin A deficiency manifests itself, depending onthe duration of deprivation in hornification of the mucous membranes(Biesalski, H. K. Stofft, E., Wellner U., Niederauer, U., and Bassler,K. H. Vitamin A and ciliated cells. I. Respiratory epithelia, "Z. F.Ernahrungswissenschaft: 25, 114-122 (1986), and McDowell E. M., KeenanK. P., Huang M.: Effect of Vitamin A deprivation on Hamster, BrachialEpithelium, Virchow's Arch. (Cell. Pathol.), 45: 197-219, 1984 andMcDowell E. M., Keenan K. P., Huang M.; Restoration of mucociliaryBrachial Epithelium following Deprivation of Vitamin A, Virchow's Arch.(Cell. Pathol.) 45: 221-240, 184), in particular of the respiratorysystem, increased susceptibility to infections and with pronounceddefect in the classic symptoms of keratinization of the conjunctiva ofthe eye up to complete destruction and blindness.

On intake of the vitamin most of the deficiency-enhanced alterations arereversible, in particular in the area of the mucous membranes. Thesetypical changes induced by the vitamin A deficiency are found in similarmanner in diseases of the mucous membranes of the respiratory system ascaused by acute and chronic bronchitis, nicotine inhalation and alsoearly cancerous changes. It has been possible to show in experimentalinvestigations on animals and on humans that such changes not induced byvitamin A deficiency could be made reversible by systemic intake of thevitamin, usually in high doses. In particular in various forms of cancerthe use of large-dose systemic vitamin A therapy is described withvarying success in the sense of preventing recidivation after primarytreatment of the tumor.

In systemic administration, to obtain a regression of the squamousmetaplasia or to prevent renewed occurrence of such changes highconcentrations must be used which in some cases lead to substantial sideeffects (cerebral pressure symptoms, liver cell metabolism disturbances,and others). In addition, there is a pronounced teratogenic effect inthe first three-month period which forbids use of large-dose therapy inpregnancy (Bauernfeind JC. (1980): The safe use of vitamin A, Thenutrition foundation, Washington D.C.).

Furthermore, an intake of vitamin A is also inadmissible inphysiological concentration in the presence of disturbances of livercell metabolism such as inflammation or cirrhosis because due to thesimultaneously disturbed protein synthesis (lack of formation of thetransport protein--RBP=retinol-binding protein) of the liver removal ofthe vitamin from the stores into which it is transferred afterabsorption is not possible and consequently further damage to the liverwould arise (Smith, F. R. and Goodman, D. S. The effect of diseases ofthe liver, thyroid, and kidneys on the transport of vitamin A in humanplasma. J. Clin. Invest.: 50, 2426 (1971); Weber, F. L., Mitchell, G.E., Powell, D. B., Reiser, B. J. and Banwell, J. G., Reversiblehepatoxicity associated with hepatic vitamin A accumulation in a proteindeficient patient. Gastroenterology: 82, 118-123 (1982)). In addition,the vitamin can only be taken up by the peripheral target tissue, suchas the respiratory epithelium, and carry out its function when it isbound to precisely this transport protein.

An object of the present invention is therefore to avoid thesedisadvantages, in particular to permit an action of the vitamin A orsimilar compounds on the mucous membranes of the urogenital tract,intestinal tract and the respiratory epithelium, the epithelia of thenose-throat cavity and preferably the epithelia of the tracheal and deepbronchial tract.

SUMMARY OF THE INVENTION

This problem is solved according to the invention by a pharmaceuticalpreparation which consists of retinoic acid and/or an ester of retinoicacid and/or an ester of retinol as active substances, which are presentin the preparation form of an aerosol. This permits in particular atopical action of the active substance on the mucous membranes.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a plot showing the distribution of retinol and retinyl estersin the body at various times after parenteral administration of retinylmargarinate.

FIGS. 2a and 2b are chromatogram traces of vitamin A derivatives inaccordance with the invention ten and 48 hours, respectively, after theinfusion of retinyl margarinate.

FIG. 3 is a bar graph representing the level of retinol in selectedtissues of animals who were deprived of Vitamin A 48 hours after theinfusion of a C₁₇ retinyl ester.

DETAILED DESCRIPTION OF THE INVENTION

In particular, according to the invention esters of retinol withphysiologically compatible carboxylic acids and esters of retinoic acidwith alcohols can be advantageously employed.

Particularly advantageous as carboxylic acids for preparing the retinolesters according to the invention are the saturated and unsaturatedfatty acids, in particular the endogenously occurring fatty acids.

Of quite particular advantage are palmitic acid and/or stearic acidand/or oleic acid and/or linoleic acid and/or linolenic acid.

Particularly advantageous alcohols to be used according to the inventionfor preparing the retinoic acid esters are monovalent and/or multivalentprimary and/or secondary and/or tertiary aliphatic and/or alicyclicand/or aromatic alcohols.

Very particularly advantageous are the aliphatic, monovalent primaryalcohols, in particular methanol and ethanol and the fatty alcohols.

Sprays and inhalates, in particular deep-action inhalates, can inparticular be used as aerosols according to the invention.

According to the invention it is achieved that the active substance isconverted to an aerosol so that the finely distributed minute activesubstance particles of the aerosol reach the place of action, forexample the mucous membrane of the respiratory system.

According to the invention aerosols or sprays mean dispersed systems ofgases with solid or liquid particles distributed therein of about 10⁻⁷to 10⁻¹ cm diameter. Consequently, with regard to the pharmaceuticalpreparation according to the invention the following distinctions are tobe made:

A) distributions of solid active substance particles in the carrier gas(dust aerosols) and

B) distributions of active-substance-containing liquid in the carriergas (mist aerosols).

The so-called mist aerosols can in turn be present as

a) distribution of the active substance dissolved in a non-aqueouslipophilic readily volatile and physiologically compatible solvent inthe carrier gas or as

b) distribution of a solution or dispersion of the active substance oran emulsion of the active substance present in liquid form in water ormixtures of water and/or water-miscible physiologically compatibleliquids in the carrier gas.

Among others, the following known technological systems are possible forthe technical execution of preparations which serve for administrationof an active substance according to the invention in the form of a sprayor aerosol distribution:

1. Pump atomizer:

By means of a piston pump mechanism in the spray head an excess pressureis generated in the atomizer vessel containing the active substancesolution or dispersion and as a result the active-substance-containingliquid is forced through the atomizer jet and thus distributed in thesurrounding air (advantageously suitable for aerosols of the type Bb).

2. Aerosol propellent gas packs:

A propellant gas ensures the excess pressure in the container from whichby opening the closure valve the suspended or dissolved active substanceis forced through the atomizer jet and distributed. A distinction ismade between:

a) Liquid gas systems:

A liquefied gas is used as propellent gas (e.g. low-boiling FCHC orpropane, butane), the system being designed as

aa) two-phase aerosol:

here the propellent gas in the pressure container is in the liquid andgaseous phase, the liquid phase simultaneously containing dissolved theactive substance and any auxiliary substances, such as additionalsolvents (advantageously suitable for aerosols of type Ba).

ab) suspension aerosol: the active substance particles are suspended insolid form in the liquid propellent phase (advantageously suitable foraerosols of type A).

ac) three-phase aerosol:

in addition to the gaseous and liquid propellent phase there is a liquidphase of the active substance solution not miscible with the propellent.In contrast to 2aa) and 2ab) in the spraying operation as long as thepackage still contains active substance phase no propellent is ejected,the latter serving in this case solely to generate the operatingpressure in the package (advantageously suitable for aerosols of typeB).

b) pressurized gas system:

in this case instead of the liquefied gas a compressed gas (e.g.nitrogen, carbon dioxide, dinitrogen monoxide, air) is used. When thevalve is of quite particular advantage are palmitic acid and/or stearicacid and/or oleic acid and/or linoleic acid and/or linolenic acid.

Particularly advantageous alcohols to be used according to the inventionfor preparing the retinoic acid esters are monovalent and/or multivalentprimary and/or secondary and/or tertiary aliphatic and/or alicyclicand/or aromatic alcohols.

Very particularly advantageous are the aliphatic, monovalent primaryalcohols, in particular methanol and ethanol and the fatty alcohols.

Sprays and inhalates, in particular deep-action inhalates, can inparticular be used as aerosols according to the invention.

According to the invention it is achieved that the active substance isconverted to an aerosol so that the finely distributed minute activesubstance particles of the aerosol reach the place of action, forexample the mucous membrane of the respiratory system.

According to the invention aerosols or sprays mean disperse systems ofgases with solid or liquid particles distributed therein of about 10⁻⁷ t10⁻¹ cm diameter. Consequently, with regard to the pharmaceuticalpreparation according to the invention the following distinctions are tobe made:

A) distributions of solid active substance particles in the carrier gas(dust aerosols) and

B) distributions of active-substance-containing liquid in the carriergas (mist aerosols).

The so-called mist aerosols can in turn be present as

a) distribution of the active substance dissolved in a non-aqueouslipophilic readily volatile and physiologically compatible solvent inthe carrier gas or as

b) distribution of a solution or dispersion of the active substance oran emulsion of the active substance present in liquid form in water ormixtures of water and/or water-miscible physiologically compatibleliquids in the carrier gas.

Among others, the following known technological systems are possible forthe technical execution of preparations which serve for administrationof an active substance according to the invention in the form of a sprayor aerosol distribution:

1. Pump atomizer:

By means of a piston pump mechanism in the spray head an excess pressureis generated in the atomizer vessel containing the active substancesolution or dispersion and as a result the active-substance-containingliquid is forced through the atomizer jet and thus distributed in thesurrounding air (advantageously suitable for aerosols of the type Bb).

2. Aerosol propellent gas packs:

A propellent gas ensures the excess pressure in the container from whichby opening the closure valve the suspended or dissolved active substanceis forced through the atomizer jet and distributed. A distinction ismade between:

a) Liquid gas systems:

A liquefied gas is used as propellent gas (e.g. low-boiling FCHC orpropane, butane), the system being designed as aa) two-phase aerosol:here the propellent gas in the pressure container is in the liquid andgaseous phase, the liquid phase simultaneously containing dissolved theactive substance and any auxiliary substances, such as additionalsolvents (advantageously suitable for aerosols of type Ba).

ab) suspension aerosol:

the active substance particles are suspended in solid form in the liquidpropellent phase (advantageously suitable for aerosols of type A).

ac) three-phase aerosol:

in addition to the gaseous and liquid propellent phase there is a liquidphase of the active substance solution not miscible with the propellent.In contrast to 2aa) and 2ab) in the spraying operation as long as thepackage still contains active substance phase no propellent is ejected,the latter serving in this case solely to generate the operatingpressure in the package (advantageously suitable for aerosols of typeB).

b) pressurized gas system:

in this case instead of the liquefied gas a compressed gas (e.g.nitrogen, carbon dioxide, dinitrogen monoxide, air) is used. When thevalve is opened the active substance phase is forced via the rise tubedipping thereinto through the atomizer valve. The propellent gas reachesthe outside only to the extent in which it is soluble in the activesubstance phase; when using a two-chamber pressurized pack however thisis not the case at all because the active substance phase is disposed ina separate plastic bag in the pressurized can and is not in directcontact with the propellent gas (advantageously suitable for aerosol oftype B).

Thus, according to the invention the pharmaceutical preparationaccording to the invention is made in that the active substance isdissolved or dispersed in a suitable nontoxic medium and said solutionor dispersion atomized to an aerosol, i.e. distributed extremely finelyin a carrier gas. This is technically possible for example in the formof aerosol propellent gas packs, pump aerosols or other devices knownper se for liquid misting and solid atomizing which in particular permitan exact individual dosage.

According to the invention the carrier gas may be part of thepharmaceutical preparation in the from of a propellent gas or a solventvaporizing on atomization, as in the case of the aerosol propellent gaspackage. The carrier gas may also however be atmospheric air which foratomizing the active-substance-containing liquid is conducted forexample as strong air stream over the opening of a capillary tube whichdips into the liquid and through which the solution is sucked up andthen dispersed in the carrier gas stream. Said carrier gas stream mayfor example be generated by a compressor integrated into the atomizerapparatus. Finally, the stationary ambient air may also be sued ascarrier gas, in which the fine droplets of theactive-substance-containing liquid are dispersed after they have beengenerated for example in the atomized jet of a pump spray, three-phaseaerosol or a two-chamber pressure pack.

Another possibility for the atomization is a mechanical dispersion ofthe liquid containing the active substance or the active substances, forexample by ultrasonic waves for example with the aid of a commerciallyavailable ultrasonic inhalator.

The active substance or the active substance combination is preferablyused in stabilized form; this is achieved for example by addition ofbutyl-hydroxyanisole (BHA) and/or butylated hydroxytoluene (BHT) orgamma-tocopherol or alpha-tocopherol or tocopherol mixtures or otherantioxidants or mixtures thereof.

The stabilization is effective irrespective of the nature of the vitaminA derivative employed.

The active substances according to the invention may be present eitheras solid particles in the carrier gas or in dissolved form in thecarrier gas.

For the case where the active substances are present in dissolved formin the carrier gas according to the invention two possibilities areprovided, i.e, an active substance solution in a nonaqueous volatilesolvent or an active substance solution in an aqueous solvent.

The solution of the active substance according to the invention maypreferably be in a nonvolatile nontoxic physiologically compatiblesolvent.

Particularly advantageous is the use of a dosage aerosol propellent gaspack. For the production thereof the active substance is first dissolvedin a toxicologically harmless inert solvent; suitable for this purposeare in particular fluorochlorohydrocarbons such as1,1,2-trichloro-1,2,2-trifluoroethane (Frigen 13) which simultaneouslyalso acts as a component of the propellent gas mixture. The solution isthen introduced into a pressure-resistant container, for example into analuminum can provided with an internal protective lacquer. Thereafter apropellent gas or propellent gas mixture consisting of liquified orcompressed gases, for example a mixture of low-boilingfluorochlorohydrocarbons, is introduced into the can under pressureafter removal of the air from said can and the can sealed with ametering valve. The components of the propellent gas are toxicologicallyharmless and inert; suitable for example are dichlorodifluoromethane(Frigen 12) or 1,2-dichloro-1,1,2,2-tetrafluoroethane (Frigen 114) orliquefied gas, such as propane, butane or dimethyl ether.

Apart from this method of the so-called pressure filling according tothe invention for example the refrigeration method or the "under the cupfilling" method are possible.

The pharmaceutical preparation prepared according to the invention inthis manner consists of:

0.01-50% by weight active substance

0-49.99% by weight solvent and

50-99.99% by weight propellent gas.

Preferably it consists of

0.01-30% by weight active substance.

0-30% by weight solvent and

40-99.99% by weight propellent gas.

In particular, preparations are suitable comprising:

0.05-25% by weight active substance

0-30% by weight solvent and

45-99.95% by weight propellent gas.

Preparations are advantageous comprising:

0.1-15% by weight active substance

0-25% by weight solvent and

60-99.9% by weight propellent gas.

Particularly advantageous are preparations comprising:

0.1-10% by weight active substance

0-25% by weight solvent and

65-99.9% by weight propellent gas.

Preparations have been found particularly advantageous which comprise:

0.1-7% by weight active substance

5-23% by weight solvent and

70-94.9% by weight propellent gas.

Also particularly advantageous is the use of preparations comprising:

0.1-5% by weight active substance

10-20% by weight solvent and

75-89.9% by weight propellent gas.

Excellent results are obtained with preparation comprising:

0.17% by weight active substance

19.83% by weight solvent and

80% by weight propellent gas.

for the administration of small individual doses of the active substanceand preparations comprising:

3.9% by weight active substance

20% by weight solvent and

76.1% by weight propellent gas.

for administrations of large individual doses of the active substance.

The active substance contents of the last two preparations relate toretinol palmitate (1.7 million IU/g). When using other active substancesor active substance mixtures the proportion in percent by weight is tobe amended so that the vitamin A activity (expressed in IU) or vitamin Aequivalent dosage (on a molar basis) contained in the respective amountof active substance remains unchanged. The resulting composition shiftsare compensated for by appropriate modification of the percentage byweight of the solvent proportion in the preparation.

The preferred discharge amount of the metering valves employed is forexample between 35 mg and 100 mg per spray shot.

Further features of the invention will be apparent from the followingdescription of nonlimiting examples of embodiment.

EXAMPLE 1

    ______________________________________    Retinol palmitate      3.9% by weight    (1.7 million IU/g),    stabilized with BHA/BHT    1,1,2-trichloro-1,2,2-trifluoroethane                           26.1% by weight    propellent gas mixture consisting of                           70.0% by weight    40% by weight dichlorodifluoromethane and    60% by weight 1,2-dichloro-1,1,2,2-    tetrafluroethane    ______________________________________

To prepare the active substance solution 13.0 kg retinol palmitate (1.7million IU/g) is dissolved in 87.0 kg trichlorotrifluoroethane. Thedissolving is carried on in a closed container with incorporatedagitator so that no evaporation of the solvent can take place. After thedissolving the liquid is filtered through a 50 μm filter and the amountof solvent evaporated when this is done replenished. For the filling theclear active substance solution is removed from a tightly sealedcontainer with a withdrawal conduit by means of a mechanical pistondosage device and introduced into the aerosol cans provided for thispurpose. The dosage device is fixed so that it always discharges thesame amount of 4.5 g per dosage stroke. Thereafter, the aerosol meteringvalve is fitted over the opening of the can and the valve undetachablysecured (crimped) on the aerosol can using automatic crimping tongs. Thecan thus sealed is now filled via an automatic propellent gas fillingapparatus through the valve with the respective propellent gas mixture.The filling amount for 4.5 g active substance solution is 10.5 gpropellent gas. The metering valve employed is advantageously designedfor a discharge of 75 mg per spray shot.

After checking the cans to ensure they are leakfree in accordance withthe technical regulations for gases (TRG 402) said cans are providedwith the spray head or a corresponding atomizer means which is fittedonto the valve ball of the metering valve. To protect the atomizer meansfrom actuation and damage the can is also provided with a suitableprotective cap or packed in a protective envelope.

EXAMPLE 2

    ______________________________________    Retinol palmitate (1.7 million IU/g)                           0.17%   by weight    stabilized with BHA/BHT    1,1,2-trichloro-1,2,2- 19.83%  by weight    trifluoroethane    propellent gas mixture consisting of                           80.0%   by weight.    40% by weight dichlorodifluoromethane and    60% by weight 1,2-dichloro-1,1,2,2-    tetrafluoroethane    ______________________________________

The metering valve employed is advantageously designed for a dischargeof 35 mg per spray shot.

EXAMPLE 3

    ______________________________________    retinol laurate, stabilized with                           0.14% by weight    alpha-tocopherol    propellent gas mixture consisting of                           99.86% by weight    40% by weight chlorotrifluoroethylene and    60% by weight dichlorofluoromethane    ______________________________________

The metering valve employed is advantageously designed for a dischargeof 35 mg per spray shot.

EXAMPLE 4

    ______________________________________    retinoic acid ethyl ester                           9.2% by weight    trichlorofluormethane  20.8% by weight    propellent gas mixture consisting of                           70% by weight    45% by weight 1,1-difluoroethane and    55% by weight 1,1-dichloro-1,2,2,2-    tetrafluorethane    ______________________________________

The metering valve employed is advantageously designed for a dischargeof 75 mg per spray shot.

EXAMPLE 5

Active substance mixture (stabilized with alpha-tocopherol) consistingof:

    ______________________________________    retinoic acid capryl ester                           1.15% by weight    retinol acetate        1.40% by weight    1,1,2-trichloro-1,2,2-trifluoroethane                           22.45% by weight    propellent gas mixture consisting of                           75.0% by weight    40% by weight 1,1-difluoroethane and    60% by weight 1,1-dichloro-1,2,2,2-    tetrafluoroethane    ______________________________________

The metering valve employed is advantageously designed for a dischargeof 75 mg per spray shot.

EXAMPLE 6

Active substance mixture (stabilized with gamma-tocopherol) consistingof:

    ______________________________________    retinol propionate  0.05% by weight    retinol oleate      0.085% by weight    trichlorofluoromethane                        19.865% by weight    propane             80.0% by weight    ______________________________________

EXAMPLE 7

    ______________________________________    retinol acetate, stabilized with                            22% by weight    gamma-tocopherol    1,1,2 trichloro-1,2,2-trifluoroethane                            22% by weight    propellent gas mixture consisting of                            56% by weight    50% 1-chloro-1,1-difluoroethane and    50% tetrafluoromethane    ______________________________________

The metering valve employed is advantageously designed for a dischargeof 35 mg per spray shot.

The preparation method for examples 2 to 6 is analogous to thatdescribed under example 1. In the case of example 3 a third of thetrichlorofluoromethane contained in the propellent gas mixture servesfor dissolving the active substance.

The active substance mist discharged by a spray shot of the meteringvalve is introduced by suitable applicators into the body cavities andonto their mucous membranes. For example, it can be introduced into themouth cavity by a mouthpiece fitted onto the spray can. From there, oninhalation after the spraying operation particles of 0.5-5 μm reach thepulmonary alveoli whilst particles of 5-30 μm are transported onto themucous membranes of the upper respiratory tract.

Further pharmaceutical preparations according to the invention are thosein which the active substances are dissolved or dispersed in aqueoussolvents. These may be referred to as "aqueous systems". "Aqueoussystems" is the term applied here to systems which contain as solventwater or mixtures of water with other physiologically compatiblesolvents.

Auxiliary substances known per se to the expert, such as emulsifiersand/or solutizers, may preferably be added to the solvent.

The active-substance-containing liquid prepared in this manner canpreferably be used

a. In the form of a three-phase liquid gas aerosol pack: the samepropellent gas systems are employed as in the two-phase aerosoldescribed above.

b. In the form of a pressurized gas aerosol: the liquid is introducedinto a spray can which is subjected to pressure by means of compressedgas (e.g. nitrogen or carbon dioxide).

c. In the form of a pump aerosol.

The active substance is preferably used already stabilized with theaforementioned antioxidants. In the case of an open system, such as thepump aerosol, in which outer air gets into the container, thepreparation is preferably conserved for example by addition of parabensor other suitable preservatives.

The aqueous active substance system which can be used in the mannerdescribed consists of:

0.01-50% by weight active substance

1-30% by weight emulsifier and/or solutizer ad 100% by weight water andcarrier gas.

Particularly suitable are systems comprising:

0.05-20% by weight active substance

5-30% by weight emulsifier and/or solutizer ad 100% by weight water andcarrier gas.

Advantageous is the use of systems comprising:

0.1-10% by weight active substance

5-25% by weight emulsifier and/or solutizer ad 100% by weight water andcarrier gas.

In particular systems comprising:

0.5-5% by weight active substance

10-25% by weight emulsifier and/or solutizer ad 100% by weight water andcarrier gas.

A nonrestrictive example for an active system to be used according tothe invention is disclosed below:

    ______________________________________    retinol palmitate oily (1 million IU/g)                         1.0% by weight    stabilized with BHA/BHT    emulsifier (Cremophor RH 40 (BASF))                         22.0% by weight    1,2-propylene glycol 2.0% by weight    water and carrier gas                         ad 100.0% by weight    ______________________________________

The pharmaceutical preparation according to the invention canadvantageously be employed for preventing and treating functionalimpairments, diseases and pathological changes in the mucous membranesof humans and animals, in particular in the respiratory epithelium, theepithelia of the nose-throat cavity, the urogenital tract and theintestinal tract.

To the functional impairments, diseases and pathological changes whichcan be influenced by the invention belong in particular cellulardifferentiation disturbances of the mucous membranes of the respiratorytract, squamous metaplasia irrespective of the genesis, neoplasticchanges, reduced activity of the ciliated epithelium, dysfunction ofmucigenous cells irrespective of the genesis.

Consequently, the pharmaceutical preparations according to the inventionare suitable inter alia for therapy or as adjuvant in the therapy ofbronchial carcinomas, acute and chronic bronchitis, acute and chronicfunctional disturbances due to impairment of the tracheobronchialepithelium following inhalation of dusts and gases damaging the mucousmembranes, bronchopulmonary dysplasia of newborn children and thecarthagena syndrome.

The necessary daily doses lies as a rule between 100 and 50,000 IUvitamin A or vitamin A equivalent dose which subject to thetoxicological acceptance as regards systemic and other side effects isadministered in 1 to 10 individual doses of 100-5,000 IU vitamin Aactivity or vitamin A equivalent dose in corresponding intervals.However, higher individual or daily doses may be administered dependingon the nature of the diseases being treated.

The invention has in particular the following advantages:

The invention ensures that the vitamin A and the other active substancesaccording to the invention in the administration form chosen accordingto the invention are absorbed into the target cells.

According to the present state of the art for obtaining a remission ofsquamous metaplasia in humans and animals high to maximum concentrationsof vitamin A must be used systemically (100,000-500,000 IU/day for up to60 days). Hitherto, it was thus possible to show only inadequately thatsquamous metaplasia which is not induced by vitamin A deficiency is alsoquantitatively returned into the mucous-secretory original epitheliumand that the metaplastically changed cell can also take up the vitamin.

The reason for this is in particular that the metaplastically changedcell no longer has at its disposal receptors for taking up the vitaminRBP complex. It was thus not to be expected that a topical intake ofvitamin A without binding protein can be accepted by the cell and usedfor metabolic purposes in order to effect in this manner a reversibilityof the squamous metaplasia.

According to the invention it was possible to show that:

1. Vitamin A, for example in the form of its long-chain fatty acidesters, can be absorbed into the cells of the respiratory mucousmembrane without mediation of receptors and/or binding proteins aftertopical administration.

2. The vitamin absorbed is converted to a metabolically activatableform.

3. The squamously metaplastically changed epithelium of the respiratorymucous membrane can be restored to its original epithelium after topicaladministration of the vitamin.

4. The dosages necessary for achieving adequate, i.e. effective localvitamin A concentrations in the respiratory epithelium when using anaerosol lie far below those necessary for systemic administration.

In this connection the following tests were carried out and will beexplained in detail with the aid of the attached FIGS. 1, 2a, 2b and 3:

The Figures show:

FIG. 1:

Distribution of retinol and retinyl esters at various times afterparenteral administration of the retinyl margarinate. Vitamin Adeficiency: in this case at the start of the test no derivatives ofretinol were present in the tissues whilst after 48 hours an appreciableenrichment can be observed in particular in the lungs and trachea whichin particular for the trachea exceeds the concentration of storedretinol esters to be measured in normal animals (rear axis).

FIGS. 2a and 2b:

Chromatogram of the investigated vitamin A derivatives 10 hours afterinfusion of the retinyl margarinate (2a) and 48 hours after infusion(2b).

FIG. 3:

Occurrence of retinol in various target tissues of animals deprived ofvitamin A 48 hours after infusion of a C 17 retinyl ester. At the startof the tests these tissues did not include any retinol so that thelatter can have penetrated the tissues only by hydrolysis of the C 17ester. In particular, no further retinol occurrence is observed in theliver and this again proves that no transesterification and separationvia the liver has taken place to a quantitatively significant extent.The detection of retinol in the cells further proves that the hydrolysisof the esters has taken place in these tissues.

Remarks on the above tests:

1)

Rats deprived of vitamin A in which no peripheral vitamin A stores couldbe detected in the respiratory epithelium were given a nonphysiological(i.e. normally not occurring but metabolizable) retinyl ester indifferent concentrations via an implanted infusion system for 6 hours.The kinetics were determined through continuous blood analyses so that48 hours after the start of the test it was possible to take organspecimens to investigate the distribution to the retinyl esters in themucous membranes of the respiratory tract but also in other tissues. Itwas found that the administered retinyl ester is absorbed in the mucousmembrane, hydrolyzed and reverse esterified so that it is present in itsmetabolizable active form (FIG. 1). This therefore means that absorptionof the vitamin A derivative without binding proteins is possible and thevitamin is therefore available to the target cells.

The topical administration of the C-17 retinyl ester by an atomizingmethod via a tracheal probe in nutritionally deficient animals led to acomparable result. The C-17 esters are absorbed in the target cells ofthe respiratory epithelium (FIG. 2a) and converted to a metabolicallyactive form (FIG. 2b) as chromatographic measurements have shown.

The use of C 17 retinyl esters in animals with a nutritional vitamin Adeficiency has several advantages:

Firstly, the nutritionally deficient animals do not have any vitamin Aoccurrences in the central and peripheral storage tissues so that ameasurement of the distribution of such esters after administration iseasily possible. Secondly, a C 17 retinyl ester does not naturally occurso that on detection in the target cells it can be assumed that thisester has thus been absorbed into the cell (see also FIG. 2a). Inaddition, with the aid of further retinyl esters occurring having fattyacids of different chain length hydrolysis and reverse esterification ofthe C 17 ester can be concluded (see also FIG. 2b).

2)

It was possible to clarify this point in the tests as described under1). The metabolically active form of the vitamin A is retinol whichafter hydrolysis of the retinyl esters can develop its effect in thetarget cells coupled to the cellularly retinol-binding protein (CRBP).This is however possible only when the vitamin is already present in thecells as ester because the absorption of the retinol through the bloodsystem after enteral absorption or liberation from the liver is possibleonly via the transport protein in the plasma (RBP). As FIG. 3 shows thetopically administered C 17 retinyl ester was absorbed into the cell,esterified after hydrolysis with palmitic acid, again hydrolyzed and isthen present in the cells as retinol (FIG. 3).

3)

In normally fed rats by chronic inhalation of cigarette smoke condensate(CSC) a squamously metaplastic change of the tracheobronchial epitheliumwas induced. This metaplastic change was particularly stronglypronounced in the bronchus of the 2nd type and is distinguished by basalcell hyperplasia and focal ciliary loss. These changes were verypronounced in all the animals (n=20) treated. The systemicadministration of 15,000 IU vitamin A as retinyl palmitate (2 times perweek) led after on the average 22 days to a complete regression of thechanges when the CSC inhalation had been stopped at the start of thevitamin A supplementation. If during the supplementation CSC inhalationwas continued the changes were reversed later (on the average after 39days) but not completely. Although the cilioneogenesis was clearlystimulated (enhanced thymidine labelling index) a few areas remained inwhich the basal cell hyperplasia was completely reversed, although therewere still a few focal ciliary losses.

In topical administration of the vitamin by an atomizer or nebulizer viaa tracheal probe with the administration twice a day each of 1000 IUvitamin A as retinyl palmitate after an average of 16 days (on ceasingthe CSC inhalation at the start of the topical administration) ascomplete remission of the squamously metaplastic changes was observedand if the CSC inhalation was continued a complete remission alsooccurred after 16 days.

This shows that the topical administration of the retinyl esters has thegreat advantage over the systemic administration that even withcontinuous irritation of the tissue the regeneration is initiated andthus the development of neoplastic changes prevented. The decisive pointis however also that far smaller concentrations were required to obtainremission and consequently systemic-toxical side effects could beavoided.

4)

High enteral vitamin A administration (>25,000 IU/day) leads to achronic hypervitaminosis A which in particular in children and pregnantwomen can lead to damage. Consequently, supplementation with thisvitamin is therapeutically useful in human beings only in subtoxic dosesof less than 25,000 IU/day. Since due to the degeneration of themembrane-bound retinol receptors the squamously metaplastically changedcell can scarcely absorb retinol from the plasma, it is dependent on theplasma retinyl esters which can reach the cell without receptors. Undernormal circumstances retinyl esters occur in plasma with normal vitaminA intake only in a very low concentration so that adequate retinyl esterplasma levels are reached only on enteral supplementation of more than100,000 IU/day. The normal concentration of on average 50 μg retinylester/g dry weight in the respiratory mucous membrane can be increasedin rats by supplementation with at least 15,000 IU vitamin A/day to 75μg/g. The plasma retinyl ester concentration is then about 50% of theretinyl concentration, corresponding in humans to a daily dose of100,000 IU vitamin A. If the vitamin is applied topically to the mucousmembrane (1000 IU vitamin A twice daily) the original concentration inthe mucous membrane rises from 50 μg/g to 110 μg/g without anappreciable increase in the plasma retinyl ester concentrationoccurring. Since the toxic side effects of high enteral vitamin A intakeare related to the increase in the plasma retinyl ester, the topicaladministration has a clear advantage of the systemic intake. Toxic sideeffects can be avoided in spite of longer application because firstlytotal concentrations for obtaining a remission are lower than withenteral intake, and an increase in the plasma retinyl esters as must beaimed at after systemic intake does not take place.

Our investigations have shown that vitamin A in the form of its retinylesters without receptors is absorbed after topical administration by thecells of the respiratory epithelium and transferred to its metabolicallyactive form. This result justified the investigation of the efficacy ofa topical administration on the reversibility of squamous metaplasia ofthe respiratory epithelium. It was possible to show that in the case ofchronic vitamin A depletion by topical administration of the aerosol.

a. the absorption into the squamously metaplastically changed cellstakes place;

b. the squamously metaplastically changed cells are restored to theiroriginal phenotype;

c. to obtain this restoration in the topical administration subtoxic,i.e. physiological, concentrations were sufficient whilst with systemicadministration only the large-dosage administration was successful.

By using a vitamin A-containing aerosol, squamous metaplasia caused byacute and chronic bronchitis, vitamin A deficiency, inhaled carcinogenicsubstances, or physical and chemical irritations, can be restored to itsoriginal phenotype. The aerosol employed has the advantage that systemiceffects, in particular of toxic nature, can be avoided.

Advantageously the invention can be applied in particular for treatingmetaplasia of the cells of the mucous membranes of the respiratorytract, in particular the squamous epithelium.

What is claimed is:
 1. A method for preventing and treating mucosaldiseases of the epithelia of the tracheobronchial tract, said epitheliabeing susceptible to treatment with vitamin A, in a human or animalsubject, said method comprising topically applying to the mucosal siteof the epithelia of a subject suffering from said disease atherapeutically effective amount of a pharmaceutical preparation in theform of an aerosol inhalate comprising at least one active substanceselected from the group consisting of an ester of retinoic acid and anester of retinol for those in need thereof, with solid or liquidparticles distributed in the formed inhalate of about 10⁻⁷ to 10⁻¹ cm indiameter.
 2. A method for preventing and treating a condition selectedfrom functional anomalies, diseases and pathological changes in themucous membranes of the respiratory epithelium, the epithelia of thetracheal tract, or the epithelia of the deep bronchial tract, saidepithelia being susceptible to treatment with vitamin A, of humans andanimals, said method comprising topically applying to said mucousmembranes of a subject suffering from said condition a therapeuticallyeffective amount of a pharmaceutical preparation in the form of anaerosol inhalate comprising at least one active substance selected fromthe group consisting of an ester of retinoic acid and an ester ofretinol for those in need thereof, with solid or liquid particlesdistributed in the formed inhalate of about 10⁻⁷ to 10⁻¹ cm in diameter.3. A method for preventing and treating a condition selected fromcellular differentiation disturbances of the mucous membranes of thetracheobronchial tract, squamous metaplasia irrespective of the genesis,neoplastic changes, restricting activity of the ciliary epithelium, anddysfunction of mucigenous cells irrespective of the genesis, said methodcomprising topically applying to said mucous membranes of a subjectsuffering from said condition a pharmaceutical preparation in the formof an aerosol inhalate comprising at least one active substance selectedfrom the group consisting of an ester of retinoic acid and an ester ofretinol for those in need thereof, with solid or liquid particlesdistributed in the formed inhalate of about 10⁻⁷ to 10⁻¹ cm in diameter.4. An adjuvant in the therapy of a condition selected from bronchialcarcinomas, acute and chronic bronchitis, acute and chronic functionaldisturbances due to reversible impairment of the tracheobronchialepithelium following inhalation of dusts and gases damaging the mucousmembranes, and bronchopulmonary dysplasia of newborn children, saidmethod comprising topically applying to a subject suffering from saidcondition a therapeutically effective amount of a pharmaceuticalpreparation in the form of an aerosol inhalate comprising at least oneactive substance selected from the group consisting of an ester ofretinoic acid and an ester of retinol for those in need thereof, withsolid or liquid particles distributed in the formed inhalate of about10⁻⁷ to 10⁻¹ cm in diameter.